p53 Signal Transduction Group, Edinburgh Cancer Research Centre, Institute of Genetics and Molecular Medicine, University of Edinburgh, Edinburgh, EH4 2XR, Scotland, UK.
J Immunol Methods. 2012 Apr 30;378(1-2):20-32. doi: 10.1016/j.jim.2012.01.013. Epub 2012 Feb 15.
The Anterior Gradient (AGR) genes AGR2 and AGR3 are part of the Protein Disulfide Isomerase (PDI) family and harbour core thioredoxin folds (CxxS motifs) that have the potential to regulate protein folding and maturation. A number of proteomics and transcriptomics screens in the fields of limb regeneration, cancer cell metastasis, pro-oncogenic oestrogen-signalling, and p53 regulation have identified AGR2 as a novel component of these signalling pathways. Curiously, despite the fact that the AGR2 and AGR3 genes are contiguous on chromosome 7p21.1-3, the AGR3 protein has rarely been identified in such OMICs screens along with AGR2 protein. Therefore there is little information on how AGR3 protein is expressed in normal and diseased states. A panel of three monoclonal antibodies was generated towards AGR3 protein for identifying novel clinical models that can be used to define whether AGR3 protein could play a positive or negative role in human cancer development. One monoclonal antibody was AGR3-specific and bound a linear epitope that could be defined using both pep-scan and phage-peptide library screening. Using this monoclonal antibody, endogenous AGR3 protein expression was shown to be cytosolic in four human ovarian cancer subtypes; serous, endometrioid, clear cell, and mucinous. Mucinous ovarian cancers produced the highest number of AGR3 positive cells. AGR3 expression is coupled to AGR2 expression only in mucinous ovarian cancers, whereas AGR3 and AGR2 expressions are uncoupled in the other three types of ovarian cancer. AGR3 expression in ovarian cancer is independent of oestrogen-receptor expression, which is distinct from the oestrogen-receptor dependent expression of AGR3 in breast cancers. Isogenic cancer cell models were created that over-express AGR3 and these demonstrated that AGR3 mediates cisplatin-resistance in mouse xenografts. These data indicate that AGR3 is over-expressed by a hormone (oestrogen-receptor α)-independent mechanism and identify a novel protein-folding associated pathway that could mediate resistance to DNA-damaging agents in human cancers.
AGR 基因 AGR2 和 AGR3 是蛋白二硫键异构酶（PDI）家族的一部分，具有潜在的调节蛋白折叠和成熟的核心硫氧还蛋白折叠（CxxS 基序）。在肢体再生、癌细胞转移、原致癌雌激素信号转导和 p53 调节等领域的许多蛋白质组学和转录组学筛选中，AGR2 被鉴定为这些信号通路的新组成部分。奇怪的是，尽管 AGR2 和 AGR3 基因在染色体 7p21.1-3 上是连续的，但在这些 OMICs 筛选中，AGR3 蛋白与 AGR2 蛋白很少被同时鉴定。因此，关于 AGR3 蛋白在正常和患病状态下如何表达的信息很少。针对 AGR3 蛋白生成了一组三种单克隆抗体，用于鉴定新的临床模型，以确定 AGR3 蛋白是否在人类癌症发展中发挥积极或消极作用。一种单克隆抗体是 AGR3 特异性的，结合了一个可以使用 pep-scan 和噬菌体肽文库筛选来定义的线性表位。使用这种单克隆抗体，显示内源性 AGR3 蛋白在四种人卵巢癌亚型中表达为细胞质；浆液性、子宫内膜样、透明细胞和黏液性。黏液性卵巢癌产生的 AGR3 阳性细胞数量最多。AGR3 表达仅与黏液性卵巢癌中的 AGR2 表达偶联，而在其他三种类型的卵巢癌中，AGR3 和 AGR2 表达不偶联。卵巢癌中 AGR3 的表达与雌激素受体表达无关，这与乳腺癌中 AGR3 的雌激素受体依赖性表达不同。创建了过度表达 AGR3 的同基因癌细胞模型，这些模型表明 AGR3 介导了小鼠异种移植中的顺铂耐药性。这些数据表明，AGR3 是通过一种激素（雌激素受体α）独立的机制过表达的，并确定了一种新的与蛋白质折叠相关的途径，该途径可能介导人类癌症中对 DNA 损伤药物的耐药性。
